Studies On Comparison Of Chemical Constituents Of Four Species Of Camellia Sect. Thea And The Central Nervous System Activity Of Theacrine, A Special Purine Alkaloid In C. Assamica Var. Kucha

Tea is the most popular beverage known to have beneficial effects on health in the world. On basis of comparison of chemical constituents of purine alkaloids and catechins in four species of Camellia Sect. Thea, the research discussed the future of development and utilization to C. ptilophylla, a special tea plant abundant in theobromine, and investigated the effects of theacrine, a peculiar purine alkaloid in C. assamica var. kucha, on central nervous system.In this paper, a RP-HPLC method was established to analyze the purine alkaloids and catechins in water extract of tea leaves. Four purine alkaloids and nine catechins in C. assamica var. kucha, C. ptilophylla, C. sinensis and C. assamica were identified and quantified at the same time. The results indicated theobromine, different from C. sinensis and C. assamica, is the major alkaloid (5.8 %) present in dry leaves of C. ptilphylla, which contains no or a small amount of caffeine. Furthermore, the HPLC method was used to detect caffeine content of 173 wild grown trees of C. ptilphylla, 29 caffeine-free tea trees obtained which could be cutting propagated and resolve the problem of commercial production of caffeine free tea.A large content substance was isolated from C. assamica var. kucha. Its structure was elucidated as theacrine (1, 3, 7, 9-tetramethyluric acid, 1) through spectroscopic analysis. Meanwhile, other 9 compounds were obtained from the extract of kucha leaves, which were caffeine (2), theobromine (3), (+)-catechin (4), (-)-epigallocatechin (5), (-)-gallocatechin (6), (-)-epigallocatechin 3-O-gallate (7), (-)-gallocatechin 3-O-gallate (8), 1, 2, 6-tri-O-galloyl-β-D-glucose (9) and gallic acid (10). Among these, compounds 4～10 were isolated from the plant for the first time.Purine alkaloids present in tea have significant effects on CNS. Caffeine, theophylline and theobromine are all widely used in clinical therapy. Even though these kinds of compounds have similar chemical structures, their effects on CNS may be much different. Compared with caffeine, the effects of theacrine on CNS have never been studied since its scarcity in nature and difficulty of artificial synthesis. Therefore, we investigated some behavioral and CNS effects of theacrine including ambulatory activity, forced swimming test, pentobarbital-induced sleep time, step-through test and Morris water maze test. The effects of theacrine on monoamine transmitters in brain of animals were also studied by animal models of restraint stress and activity stress. By comparison with caffeine and theobromine, the effects of theacrine on CNS could be illustrated.In the ambulatory activity and forced swimming tests, caffeine (10-30 mg/kg) could produce a remarkable increase in activity, but theobromine (10 mg/kg) responded with a reversed effect compared with caffeine. Theacrine at the dose of 10 mg/kg also showed some weakly sedative effect, whereas did not process significant effect at 30 mg/kg. In test of pentobarbital induced sleep time, theacrine at 10 or 30 mg/kg could reliably prolong the sleep time of mice in either normal or acute and chronic stress condition, whereas caffeine and theobromine increased the arousal level of mice at the same time. The step-through task and Morris water maze test results revealed that theacrine, caffeine and theobromine all could improve the learning and momory abilities.The results of restraint stress suggested that the levels of monoamine transmitters in brain of mice generally increased after restraint stress. Three purine alkaloids could increase NE, 5-HT and 5-HIAA levels in brain cortex and NE in hypothalamus in both normal and stress mice, but the action of theacrine and theobromine were much weaker than caffeine at the same dose. The activity stress paradigm could make rats have abnormal behaviors such as anorexia nervosa and compulsive behavior similar to those observed in humans. Except for 30 mg/kg caffeine, all three purine alkaloids could ameliorate the abnormal behaviors of activity stress rats. Neurochemical results showed that administration of three purine alkaloids could increase 5-HT levels in brain of activity stress rats which may account for their protective effect on behavior of stress rats. The difference of effects of three alkaloids on central nonoamine transmitters and the mechanisms involved still required further study.The chemical and bioactive studies in this academic dissertation confirmed caffeine-free characters of C. ptilophylla, and firstly revealed effects of theacrine obtained from C. assamica var. kucha on central nervous system. Our findings will provide faithful theoretical and experimental evidences for research and development of C. ptilophylla and C. assamica var. kucha in future, and also inaugurate a new way to explore systemically the other plants' resources of Camellia Sect. Thea.